Recombinant baculovirus with surface displaying African swine fever virus antigen P30 protein, preparation method and application thereof

Abstract

The invention discloses a recombinant baculovirus for displaying African swine fever virus antigen P30 protein on the surface as well as a preparation method and application of the recombinant baculovirus. A gene sequence of P30 is inserted between an N-terminal signal peptide SP sequence of baculovirus envelope glycoprotein gp64 and a fusion structural domain TMD to construct a fusion gene gp64-Fp30, then the fusion gene gp64-Fp30 is connected with a recombinant vector pFPG to construct a recombinant expression vector pFPG-gp64-Fp30, then the pFPG-gp64-Fp30 is converted into DH10Bac cells to obtain recombinant Bacmid plasmids, finally, the recombinant Bacmid plasmids are transfected into Sf9 cells, and the recombinant baculovirus envelope glycoprotein gp64 fusion protein is obtained. The recombinant baculovirus with the surface displaying the African swine fever virus antigen P30 protein is obtained. The recombinant P30 protein produced by using the recombinant baculovirus has a conformation similar to that of natural protein, can be induced to generate a serum antibody with excellent sensitivity and specificity, can also realize efficient and rapid expression of the P30 antigen, and has a wide application prospect in preparation of ASFV diagnostic reagents or ASF vaccines.

Description

technical field [0001] The invention belongs to the field of recombinant protein diagnostic reagents and vaccines, and relates to a recombinant baculovirus displaying African swine fever virus antigen P30 protein on its surface, a preparation method and application thereof. Background technique [0002] African swine fever (ASF) is a highly contagious and fatal acute febrile infectious disease caused by African swine fever virus (ASFV) infecting domestic pigs and wild boars. In 1921, ASF was first reported in Kenya, Africa. Since the disease was difficult to control, it has spread to many countries in the world after nearly a century. The World Organization for Animal Health (OIE) listed it as a legally notifiable animal disease. [0003] The inner envelope protein P30 encoded by the CP204L gene of ASFV has a protein size of 23.6kD, which is continuously expressed in the early stage of infection and participates in the virus invasion of host cells. Previous studies have sho...

AI Technical Summary

Problems solved by technology

[0005] In recent years, the P30 protein antigen-induced serum antibody produced by the P30 recombinant vector constructed by the traditional pFastBac1 vector has strong specificity, but insufficient sensitivity, and the non-combined promoter function limits the expression of foreign proteins ([3 ] Xu Lihui, Chi Lichao, Wang Yujia, et al. Eukaryotic expression of African swine fever virus P30 protein and establishment of indirect ELISA antibody detection method[J]. Chinese Journal of Preventive Veterinary Medicine, 2021,43(04):382-387.)

The monoclonal antibody produced by the ASFVP30 recombinant vector constructed by Yu Haoyang et al. has good sensitivity and specificity, but its screening preparation and purification steps are cumbersome and the cycle is long, which increases the cost of detection reagents or vaccine production ([4] Yu Hao Yang, Wang Caixia, Wu Shaoqiang, et al. Preparation of African swine fever virus p30 protein monoclonal antibody and establishment of blocking ELISA detection method[J / OL]. Chinese Veterinary Science: 1-10[2021-12-02].https: https: / / doi.org / 10.16656 / j.issn.1673-4696.2022.0001.)

Images